Enantioseparation and stacking of Cyanobenz[f]isoindole-amino acids by reverse polarity capillary electrophoresis and sulfated beta-cyclodextrin.

Anal Chem 79(2):736-43 (2007) PMID 17222044

A capillary electrophoresis method with laser-induced fluorescence detection for the chiral separation of cyanobenz[f]isoindole (CBI) derivatives of amino acids was developed and optimized. The enantioseparations are accomplished with sulfated beta-CD (S-beta-CD) as chiral selector at low pH and reverse polarity. BGE conditions were optimized for CBI-serine and then applied to other CBI-amino acids. Baseline resolution of 13 CBI-amino acids was achieved using a single BGE formulation of 2 wt % S-beta-CD in 25 mM phosphate buffer at pH 2.00 and a voltage of -30 kV. pH is the most critical BGE parameter affecting resolution. At 2 wt % S-beta-CD, CBI-serine enantiomers are baseline-resolved at pH 2.00 but no resolution is obtained at pH 3.00. l-Glutamate, l-aspartate and d-serine are simultaneously quantified in the microdialysate of an arctic ground squirrel to illustrate the application to biological samples. Dilute solutions of the CBI-amino acids in water can be stacked by hydrodynamic injection with a 100-fold improvement in signal-to-noise ratio without loss of chiral resolution. The stacking is proposed to consist of field-amplified migration, pH-mediated stacking, and sweeping by S-beta-CD. The limit of detections for CBI-dl-serine and CBI-dl-glutamate are determined as 0.20 and 0.30 nM, respectively. The stacking method was not applicable to the high ionic strength microdialysates.